A universal primer mixture for sequence determination at the 3' ends of cDNAs.
We have devised a universal primer which can be used to sequence the 3'-ends of cloned cDNAs containing a polyA tail. The primer consists of an equimolar mixture of three primers: 20 T nucleotides followed by either an A, C, or G nucleotide (5'----3'). With this primer mixture and the dideoxynucleotide chain termination method, we determined the 3'-terminal sequence of human beta-actin cDNA in an Okayama-Berg vector, in four parallel sets of reactions containing either a single primer (T20G, T20C, or T20A) or an equimolar mixture of all three primers. Priming with both T20A and the triple mixture gave clearly readable results that agree with the known sequence of the human beta-actin gene, and we have applied this method successfully to several other cDNAs in the Okayama-Berg expression vector. Use of this universal primer mixture facilitates determination of sequences at the 3'-ends of cDNAs while by-passing the polyA tail region.
Thweatt
R
R
Department of Medicine, University of Arkansas for Medical Sciences, Little Rock.
Goldstein
S
S
Shmookler Reis
R J
RJ
D000199:Actins / Q000235:genetics; D004270:DNA, Circular / Q000737:chemistry*; D006801:Humans; D009710:Nucleotide Mapping; D009711:Nucleotides; D010957:Plasmids; D011061:Poly A / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis
D000199:Actins; D004270:DNA, Circular; D009711:Nucleotides; D012333:RNA, Messenger; D011061:Poly A